Genetic analysis of a β-1,3/1,4-glucan utilization gene cluster in <i>Segatella copri</i> JCM 13464<sup>T</sup>

Barley β-glucan (BGL), a dietary fiber composed of β1,3- and β1,4-linked glucose units, confers various health benefits, including anti-diabetic effects. A previous study reported that the anti-diabetic effect of BGL was associated with gut bacteria, particularly <i>Segatella copri</i>. Recently, a study using recombinant proteins revealed the biochemical characteristics of proteins encoded by the polysaccharide utilization locus 4 (PUL4), which is implicated in BGL assimilation. However, the precise physiological roles of PUL4 remain unclear. In this study, we used gene disruption in <i>S. copri</i> JCM 13464^T (=DSM 18205^T; also known as <i>Prevotella copri</i> CB7) to investigate the physiological functions of PUL4 under BGL-supplemented conditions. Deletion of <i>pul4</i> significantly reduced bacterial growth, as well as acetic and succinic acid production, indicating that PUL4 is essential for efficient BGL assimilation and key metabolite generation. Moreover, although PUL4 contributed to BGL and lichenan utilization, cello-oligosaccharide assimilation did not require PUL4, indicating the presence of additional metabolic systems in <i>S. copri</i> JCM 13464^T. Strain comparisons showed that one of the four <i>S. copri</i> strains assimilated BGL despite lacking PUL4, implying that some strains may possess alternative BGL-degrading loci other than PUL4. These findings provide direct evidence that PUL4 is an indispensable gene cluster for BGL assimilation by <i>S. copri</i> JCM 13464^T. Because PUL4 enhances the biomass yield on BGL, it likewise boosts total acetate and succinate formation, potentially generating health benefits.